Hydrophobic polycationic coatings disinfect poliovirus and rotavirus solutions

Coating surfaces with N-alkylated polyethylenimines (PEIs), namely branched N,N-hexyl,methyl-PEI via covalent attachment to glass or linear N,N-dodecyl,methyl-PEI by physical deposition ("painting") onto polyethylene, enables the resultant materials to quickly and efficiently disinfect aqueous solutions of (non-enveloped) poliovirus and rotavirus.     

Highly efficient protein separations in high-performance capillary electrophoresis,using hydrophilic coatings on polysiloxane-bonded columns

Three methods for preparing hydrophilic coatings on polysiloxane bonded CElect H-type capillary electrophoresis columns have been shown. The polyalkylsiloxane-bonded phase is the first coating layer on the capillary surface, and nonionic surfactant, hydrophilic polymer, or polymer surfactant, adsorbed onto this first layer through hydrophobic interactions, forms the second coating layer. The resultant capillary surfaces are inert, hydrophilic, and suitable for highly efficient protein separations. The effectiveness and applicability of these capillary surface modification methods were tested for the separations of a variety of proteins over a wide range of buffer pH values under different capillary electrophoretic operation modes.     

Low molecular weight polyethylenimines linked by beta-cyclodextrin for gene transfer into the nervous system

BACKGROUND: Polyethylenimines (PEIs) with high molecular weights are effective nonviral gene delivery vectors. However, the in vivo use of these PEIs can be hampered by their cellular toxicity. In the present study we developed and tested a new PEI polymer synthesized by linking less toxic, low molecular weight (MW) PEIs with a commonly used, biocompatible drug carrier, beta-cyclodextrin (CyD). METHODS AND RESULTS: The terminal CyD hydroxyl groups were activated by 1,1'-carbonyldiimidazole. Each activated CyD then linked two branched PEI molecules with MW of 600 Da to form a CyD-containing polymer with MW of 61 kDa, in which CyD served as a part of the backbone. The PEI-CyD polymer developed was soluble in water and biodegradable. In cell viability assays with sensitive neurons, the polymer performed similarly to low-MW PEIs and displayed much lower cellular cytotoxicity compared to PEI 25 kDa. The gene delivery efficiency of the polymer was comparable to, and at higher polymer/DNA ratios even higher than, that offered by PEI 25 kDa in neural cells. Attractively, intrathecal injection of plasmid DNA complexed by the polymer into the rat spinal cord provided levels of gene expression close to that offered by PEI 25 kDa. CONCLUSIONS: The polymer reported in the current study displayed improved biocompatibility over non-degradable PEI 25 kDa and mediated gene transfection in cultured neurons and in the central nervous system effectively. The new polymer would be worth exploring further as an in vivo delivery system of therapeutic genetic materials for gene therapy of neurological disorders.     

Biomolecules in multilayer film for antimicrobial and easy-cleaning stainless steel surface applications

Microorganisms are able to attach to, grow on, and ultimately form biofilms on a large variety of surfaces, such as industrial equipment, food contact surfaces, medical implants, prostheses and operating rooms. Once organized into biofilms, bacteria are difficult to remove and kill, which increases the risk of cross-contamination and infection. One way to address the problem may thus be to develop antibacterial, anti-adhesion, ‘easy cleaning’ surfaces. In this study, stainless steel (SS) surfaces with antibacterial properties were created by embedding several antimicrobial peptides in a multilayer film architecture. The biocidal effect of these surfaces was demonstrated against both Gram-positive and Gram-negative bacteria according to two ISO tests. Also, coating SS surfaces with either mucin or heparin led to a reduction of S. epidermidis adhesion of almost 95% vs the bare substratum. Finally, by combining both antibacterial and anti-adhesion biomolecules in the same multilayer film, SS surfaces with better cleanability were produced. This surface coating property may help to delay the buildup of a dead bacterial layer which is known to progressively reduce exposure of the coating, leading to an undesirable decrease in the antibacterial effect of the surface.     

Mechanism of bactericidal and fungicidal activities of textiles covalently modified with alkylated polyethylenimine

Our previous studies have led to a novel "nonrelease" approach to making materials bactericidal by covalently attaching certain moderately hydrophobic polycations to their surfaces. In the present work, this strategy is extended beyond the heretofore-used nonporous materials to include common woven textiles (cotton, wool, nylon, and polyester). Pieces of such cloths derivatized with N-hexylated+methylated high-molecular-weight polyethylenimine (PEI) are strongly bactericidal against several airborne Gram-positive and Gram-negative bacteria. In contrast, the immobilized and N-alkylated PEIs of low molecular weight have only a weak, if any, bactericidal activity. These findings support a mechanism of the antibacterial action whereby high-molecular-weight and hydrophobic polycationic chains penetrate bacterial cell membranes/walls and fatally damage them. The bactericidal textiles prepared herein are lethal not only to pathogenic bacteria but to fungi as well.     

Use of a quartz crystal microbalance to investigate the antiadhesive potential of N-acetyl-L-cysteine

The reduction of bacterial biofilm formation on stainless steel surfaces by N-acetyl-L-cysteine (NAC) is attributed to effects on bacterial growth and polysaccharide production, as well as an increase in the wettability of steel surfaces. In this report, we show that NAC-coated stainless steel and polystyrene surfaces affect both the initial adhesion of Bacillus cereus and Bacillus subtilis and the viscoelastic properties of the interaction between the adhered bacteria and the surface. A quartz crystal microbalance with dissipation was shown to be a powerful and sensitive technique for investigating changes in the applied NAC coating for initial cell surface interactions of bacteria. The kinetics of frequency and dissipation shifts were dependent on the bacteria, the life cycle stage of the bacteria, and the surface. We found that exponentially grown cells gave rise to a positive frequency shift as long as their cell surface hydrophobicity was zero. Furthermore, when the characteristics of binding between the cell and the surface for different growth phases were compared, the rigidity increased from exponentially grown cells to starved cells. There was a trend in which an increase in the viscoelastic properties of the interaction, caused by the NAC coating on stainless steel, resulted in a reduction in irreversibly adhered cells. Interestingly, for B. cereus that adhered to polystyrene, the viscoelastic properties decreased, while there was a reduction in adhered cells, regardless of the life cycle stage. Altogether, NAC coating on surfaces was often effective and could both decrease the initial adhesion and increase the detachment of adhered cells and spores. The most effective reduction was found for B. cereus spores, for which the decrease was caused by a combination of these two parameters.     

Bacterial adhesion to a model surface with self-generated protection coating of mucin via jacalin

Using a mechanism of `self-generation', polymer surfaces were coated with ocular mucin-type glycoproteins that were extracted from tear fluid and immobilized through specific interaction with a lectin, jacalin. Separately, jacalin affinity chromatography of tear fluid showed the main retained components had molecular weights higher than 200 kDa. In evaluations of bacterial adhesion, a model surface with jacalin-immobilized ocular mucins took up a significantly smaller number of adhered Staphylococcus epidermidis (0.041×10⁶ cells cm^–2) than a bare surface of the same polymer (1.202×10⁶ cells cm^–2). The lectin-mediated ocular mucin coating reduced the bacteria uptake by about 95% showing that the presence of mucin on surfaces may afford a general protection against bacterial colonization.     

Proving the antimicrobial spectrum of an amphoteric surfactant-sol-gel coating: a food-borne pathogen study

An antimicrobial coating was evaluated in this work for its antimicrobial efficacy against common food-borne pathogens. Dodecyl-di(aminoethyl)-glycine, an organic disinfectant, was immobilized in a silicon oxide matrix to generate thin films over surfaces by means of the sol-gel process. Tetraethoxysilane was used as the polymeric precursor. No alteration of optical transparency on the covered surfaces was observed. Topographic images obtained with atomic force microscopy showed a homogeneous film with no additional roughness added by the polymer to the surface. The attenuated total reflectance-Fourier transform infrared spectral data showed the presence of dodecyl-di(aminoethyl)-glycine in the silicon oxide network after a normal cleaning procedure. The antimicrobial efficacy test was performed by exposing coated slides to suspensions of common food-borne pathogens: Escherichia coli, Staphyloccocus aureus, E. coli O157:H7, Salmonella typhi, S. cholerasuiss, Listeria innocua and L. monocytogenes. The coating activity was not only bacteriostatic but also bactericidal. The percent reduction of viable microorganism exposure over 24 h to the coated surface ranged between 99.5%, for the more resistant gram-positive bacteria, and over 99.999%, for most gram-negative bacteria. The silicon matrix itself did not account for any reduction of viable microbial, even more an increase was observed.     

The role of microbial biofilms in deterioration of space station candidate materials

Formation of microbial biofilms on surfaces of a wide range of materials being considered as candidates for use on the International Space Station was investigated. The materials included a fibre-reinforced polymeric composite, an adhesive sealant, a polyimide insulation foam, teflon cable insulation, titanium, and an aliphatic polyurethane coating. They were exposed to a natural mixed population of bacteria under controlled conditions of temperature and relative humidity (RH). Biofilms formed on the surfaces of the materials at a wide range of temperatures and RHs. The biofilm population was dominated by Pseudomonas aeruginosa, Ochrobactrum anthropi, Alcaligenes denitrificans, Xanthomonas maltophila, and Vibrio harveyi. The biocide, diiodomethyl-p-tolyl sulfone, impregnated in the polyurethane coating, was ineffective against microbial colonization and growth. Degradation of the polyurethane coatings was monitored with electrochemical impedance spectroscopy (EIS). The impedance spectra indicated that microbial degradation of the coating occurred in several stages. The initial decreases in impedance were due to the transport of water and solutes into the polymeric matrices. Further decreases were a result of polymer degradation by microorganisms. Our data showed that these candidate materials for space application are susceptible to biofilm formation and subsequent degradation. Our study suggests that candidate materials for use in space missions need to be carefully evaluated for their susceptibility to microbial biofilm formation and biodegradation.     

Computational entropy estimation of linear polyether-modified surfaces and correlation with protein resistant properties of such surfaces

The non-specific adsorption of proteins on surfaces is a well-known and mostly undesirable phenomena, which is reduced by a surface coating with the linear polyether poly(ethylene glycol) (PEG) as the current benchmark material. However, the molecular mechanism of protein-resistant surfaces is still not fully understood. Two main hypotheses are generally applied. The first one is steric repulsion of the highly flexible tethered polymer chains, leading to an entropic penalty by adsorption of proteins due to the reduction in polymer chain mobility. The second one argues with well-hydrated polymer chains generating a repulsive interfacial water layer. In this article, we compare the three different protein-resistant polyether structures PEG, linear polyglycerol (LPG(OH)) and linear poly(methyl glycerol) (LPG(OMe)) to get new insights into the molecular mechanism behind protein resistance. In a theoretical approach, we apply an entropy estimator that assesses the conformational states of the tethered polyethers from MD simulations. It reveals the entropy differences between these polyethers to be in the order PEG>LPG(OH) > LPG(OMe). Moreover, experiments on fibrinogen adsorption of these surfaces via surface plasmon resonance spectroscopy are performed and correlated with the theoretical studies. We find that protein resistant properties of surfaces are likely to arise from an interplay of different factors.     

Doubly biomimetic catecholic phosphorylcholine copolymer: a platform strategy for fabricating antifouling surfaces

A doubly biomimetic PMNC polymer bearing cell antifouling phosphorylcholine and mussel adhesive protein catechol groups is synthesized. The polymer can be deposited onto a variety of substrates by dip-coating in an aqueous solution, adhering to surfaces via the catechol functional group while at the same time forming a cell outer membrane mimetic antifouling surface. Contact angle, ATR-FTIR and XPS measurements confirm polymer coating formation on a variety of inorganic and organic substrates. BSA and bovine plasma fibrinogen protein adsorption on PMNC coated surfaces are reduced significantly compared to unmodified substrates, and platelet adhesion from human serum onto the PMNC coated substrate surfaces is highly suppressed in this study.     

From multifunctionalized poly(ethylene imine)s toward antimicrobial coatings

Primary amine groups of branched poly(ethylene imine) (PEI) were functionalized with quaternary ammonium groups, alkyl chains of different length, allylic and benzylic groups in a one-step reaction, using a carbonate coupler. The structure of the obtained amphiphilic polymers was determined by means of 1H and 13C NMR spectroscopy. Depending on their hydrophilic/hydrophobic balance, the obtained polymers can be used as water-soluble disinfectants and for antimicrobial coating materials. The bactericidal properties of some of the amphiphilic polymers against Gram-negative and Gram-positive bacteria were investigated. Minimal inhibitory concentrations (log 4 reduction of bacterial growth) against Escherichia coli and Bacillus subtilis were determined in the range of 0.3-0.4 mg/mL and 0.03-0.04 mg/mL for water-soluble polymers. Glass slides coated with functionalized PEIs showed a reduction of colony forming units of at least 95%, at best 99.9%, against E. coli and B. subtilis.     

Lysozyme for capture of microorganisms on protein biochips

Lysozyme placed on the SiO₂ surfaces that have previously been derivatized with C₁₈ coating will capture both Escherichia coli and Listeria monocytogenes cells from PBS buffer at pH 7.2. This phenomenon is of significance for the design and fabrication of protein biochips that are designed to capture bacteria from buffer or water so that these can be further interrogated with respect to possible pathogenicity. Fluorescent microscopy shows that two types of bacteria (gram-negative E. coli and gram-positive Listeria spp.) will be adsorbed by lysozyme placed on the surface of the biochip but that strong adsorption of the bacteria is reduced but not eliminated when Tween 20 is present (at 0.5%) in the PBS buffer in which the cells are suspended. In comparison, Tween 20 and Bovine Serum Albumin (BSA) almost completely block adsorption of these bacteria on C₁₈ coated surfaces. The combination of a lysozyme surface with Tween 20 gives a greater degree of adsorption of L. monocytogenes than E. coli, and hence suggests selectivity for the more hydrophobic E. coli may be reduced by the Tween 20. This paper presents protocols for preparing protein-coated, SiO₂ surfaces and the effect of buffer containing Tween 20 on adsorption of bacteria by SiO₂ surfaces coated with C₁₈ to which BSA, lysozyme or C11E9 antibody is immobilized at pH 7.2 and ambient temperature.     

Thermoresponsive poly(N-isopropylacrylamide)-based block copolymer coating for optimizing cell sheet fabrication

Thermoresponsive surfaces are prepared via a spin-coating method with a block copolymer consisting of poly(N-isopropylacrylamide) (PIPAAm) and poly(butyl methacrylate) (PBMA) on polystyrene surfaces. The PBMA block suppresses the removal of deposited PIPAAm-based polymers from the surface. The polymer coating affects the temperature-dependent cellular behavior of the surfaces with respect to protein adsorption. By adjusting layer thicknesses, PBMA-b-PIPAAm-coated surfaces are optimized to regulate the adhesion/detachment of cells by temperature changes. Thus, thermoresponsive polymer-coated surfaces are able to harvest contiguous cell sheets with their basal extracellular matrix proteins.     

Antibacterial isoeugenol coating on stainless steel and polyethylene surfaces prevents biofilm growth

Aims

Pathogenic bacteria can spread between individuals or between food items via the surfaces they share. Limiting the survival of pathogens on surfaces, therefore, presents an opportunity to limit at least one route of how pathogens spread. In this study, we propose that a simple coating with the essential oil isoeugenol can be used to circumvent the problem of bacterial transfer via surfaces.

Methods and Results

Two commonly used materials, stainless steel and polyethylene, were coated by physical adsorption, and the coatings were characterized by Raman spectroscopy, atomic force microscopy and water contact angle measurements. We quantified and visualized the colonization of coated and uncoated surfaces by three bacteria: Staphylococcus aureus, Listeria monocytogenes and Pseudomonas fluorescens. No viable cells were detected on surfaces coated with isoeugenol.

Conclusions

The isoeugenol coating prepared with simple adsorption proved effective in preventing biofilm formation on stainless steel and polyethylene surfaces. The result was caused by the antibacterial effect of isoeugenol, as the coating did not diminish the adhesive properties of the surface.

Significance and Impact of the Study

Our study demonstrates that a simple isoeugenol coating can prevent biofilm formation of S. aureus, L. monocytogenes and P. fluorescens on two commonly used surfaces.     

Algal antifouling and fouling-release properties of metal surfaces coated with a polymer inspired by marine mussels

The marine antifouling and fouling-release performance of titanium surfaces coated with a bio-inspired polymer was investigated. The polymer consisted of methoxy-terminated poly(ethylene glycol) (mPEG) conjugated to the adhesive amino acid l-3,4-dihydroxyphenylalanine (DOPA) and was chosen based on its successful resistance to protein and mammalian cell fouling. Biofouling assays for the settlement and release of the diatom Navicula perminuta and settlement, growth and release of zoospores and sporelings (young plants) of the green alga Ulva linza were carried out. Results were compared to glass, a poly(dimethylsiloxane) elastomer (Silastic T2) and uncoated Ti. The mPEG-DOPA3 modified Ti surfaces exhibited a substantial decrease in attachment of both cells of N. perminuta and zoospores of U. linza as well as the highest detachment of attached cells under flow compared to control surfaces. The superior performance of this polymer over a standard silicone fouling-release coating in diatom assays and approximately equivalent performance in zoospore assays suggests that this bio-inspired polymer may be effective in marine antifouling and fouling-release applications.     

Molecular dynamics simulations of PEI mediated DNA aggregation

Understanding the molecular mechanism of polycation induced DNA aggregation and condensation is important for optimal design of gene delivery carriers. In this work, we performed a series of all-atom molecular dynamics (MD) simulations to investigate polyethylenimine (PEI) mediated DNA aggregation. We found that PEIs condense DNA through two mechanisms: polyion bridging and electrostatic screening of the DNA charges. At PEI/DNA charge ratio >1, PEIs can completely neutralize DNAs at a short distance (～12 ? from the C1' atoms), and this distance is found to be insensitive to the exact value of the charge ratio. When excess PEIs are added to a formed DNA-PEI aggregate, they are found to bind to the aggregate and increase its cationic charge. The added PEIs can also replace the PEIs previously bound to the aggregate. The excess PEIs, however, do not change the spacing of the DNAs in the aggregates. Our simulation results shed light on the mechanisms of PEI, and more generally polycation, mediated DNA aggregation and condensation.     

Novel Concepts for the Design and Manufacture of Solid Supports for Synthesis of Peptides

Two novel concepts for the design and manufacture of polymer supports for solid phase synthesis of peptides are described. The first concept involves the encapsulation of polymers within the hole of short pieces of capillary tubing often referred to as seed beads. This provides a rigid exo-skeleton for the support of soft polymer gel and other mechanically fragile polymer based matrices. The rigidity of the support provides a polymeric media that is particularly suited to continuous flow based peptide synthesis. The second concept complements this by providing an inexpensive approach to the preparation of spherical polymer particles by coating commercially available impervious hollow glass microspheres with polymer. The added advantage of this approach lies in the buoyancy of the resultant polymer particles, which facilitates handling on a large scale.     

Effects of Quaternary Ammonium Silane Coatings on Mixed Fungal and Bacterial Biofilms on Tracheoesophageal Shunt Prostheses

Two quaternary ammonium silanes (QAS) were used to coat silicone rubber tracheoesophageal shunt prostheses, yielding a positively charged surface. One QAS coating [(trimethoxysilyl)-propyldimethyloctadecylammonium chloride] was applied through chemical bonding, while the other coating, Biocidal ZF, was sprayed onto the silicone rubber surface. The sprayed coating lost its stability within an hour, while the chemically bonded coating appeared stable. Upon incubation in an artificial throat model, allowing simultaneous adhesion and growth of yeast and bacteria, all coated prostheses showed significant reductions in the numbers of viable yeast (to 12% to 16%) and bacteria (to 27% to 36%) compared with those for silicone rubber controls, as confirmed using confocal laser scanning microscopy after live/dead staining of the biofilms. In situ hybridization with fluorescently labeled oligonucleotide probes showed that yeasts expressed hyphae on the untreated and Biocidal ZF-coated prostheses but not on the QAS-coated prostheses. Whether this is a result of the positive QAS coating or is due to the reduced number of bacteria is currently unknown. In summary, this is the first report on the inhibitory effects of positively charged coatings on the viability of yeasts and bacteria in mixed biofilms. Although the study initially aimed at reducing voice prosthetic biofilms, its relevance extends to all biomedical and environmental surfaces where mixed biofilms develop and present a problem.     

Effects of quaternary ammonium silane coatings on mixed fungal and bacterial biofilms on tracheoesophageal shunt prostheses

Two quaternary ammonium silanes (QAS) were used to coat silicone rubber tracheoesophageal shunt prostheses, yielding a positively charged surface. One QAS coating [(trimethoxysilyl)-propyldimethyloctadecylammonium chloride] was applied through chemical bonding, while the other coating, Biocidal ZF, was sprayed onto the silicone rubber surface. The sprayed coating lost its stability within an hour, while the chemically bonded coating appeared stable. Upon incubation in an artificial throat model, allowing simultaneous adhesion and growth of yeast and bacteria, all coated prostheses showed significant reductions in the numbers of viable yeast (to 12% to 16%) and bacteria (to 27% to 36%) compared with those for silicone rubber controls, as confirmed using confocal laser scanning microscopy after live/dead staining of the biofilms. In situ hybridization with fluorescently labeled oligonucleotide probes showed that yeasts expressed hyphae on the untreated and Biocidal ZF-coated prostheses but not on the QAS-coated prostheses. Whether this is a result of the positive QAS coating or is due to the reduced number of bacteria is currently unknown. In summary, this is the first report on the inhibitory effects of positively charged coatings on the viability of yeasts and bacteria in mixed biofilms. Although the study initially aimed at reducing voice prosthetic biofilms, its relevance extends to all biomedical and environmental surfaces where mixed biofilms develop and present a problem.